Opioids are invaluable in pain management. Unfortunately, chronic opioid administration can lead to numerous adverse consequences such as a progressive decline in analgesic efficacy (tolerance), insurmountable pain and addiction. Thus, the extremely high therapeutic value of opioids is diminished by the detrimental impact of chronic opioid administration and abuse liability and can inflict enormous emotional and economic cost to individuals, families and society. Unfortunately, knowledge concerning the neurobiological mechanism associated with adverse consequences of chronic opioid administration has not produced an intervention or alternative treatment medication. A new approach is required to discover alternative treatment strategies. Our recent collaborative investigations have revealed a major role for canonical pathways involved in neuroplasticity and the discovery of microRNA (miRNA) involvement in morphine analgesic tolerance and drug self-administration. MiRNA expression is a strong candidate for coordinating the complex response to chronic drug exposure due to their network-like post-transcriptional effects on neuronal differentiation and dendritic architecture. Discovering the role of specific miRNA involvement in response to opioid self-administration may open the door to novel therapeutic strategies. The purpose of this CEBRA application is to define the role of miRNAs in self-administration using two novel strategies: 1) Genetically engineered mice with cell type-specific conditional knockdown of the rate- limiting miRNA processing enzyme, Dicer1, and 2) miRNA and mRNA expression profiling to discover miRNA:mRNA regulatory pairings involved in neuroadaptive changes. Our hypothesis is that escalated drug intake and the development of the self-administration habit is caused by a coordinated change in expression of select miRNAs. The following aims are proposed: Specific Aim 1: The purpose of this aim is to generate Cre-loxP animal models to enable timed and specific Dicer1 knockdown in dopaminergic and GA- BAergic neurons (using the Slc6a3 and Gad2 promotors, respectively) and phenotype self-administration behavior (and subsequent brain tissue) in a manner that can discriminate between active reinforcement and passive drug exposure. The results of this aim will provide valuable insight into miRNA's essential role, the cell- types involved and provide a phenotypic anchor for use in Aim 2. Specific Aim 2: Regional miRNA and mRNA expression profiles will be determined in multiple brain regions following morphine self-administration. The systematic process of expression profiling genotype-dependent behavioral responses to morphine and trait-specific associations/correlations with miRNA and mRNA expression will identify candidates specifically connected to self-administration. Computational methods will help prioritize miRNA:mRNA regulatory pairings and provide a functional framework to place the high-value pairs into targetable biological pathways. The multi- tiered, multi-disciplinary approach (behavior, molecular, bioinformatics) is designed to probe an unchartered layer of the genetic architecture (miRNA) to discover novel molecular targets for therapeutic intervention.